The directed evolution of NDM-1

beta-Lactam antibiotics are among the most frequently prescribed therapeutic agents. A common mechanism of resistance toward beta-lactam antibiotics is the production of beta-lactamases. These enzymes are capable of hydrolyzing the beta-lactam bond, rendering the drug inactive. Among the four described classes, the metallo- beta-lactamases (MBLs, class B) employ one or two zinc ions in the active site for catalysis. One of the three most clinically relevant MBLs is New Delhi Metallo- beta-Lactamase (NDM-1). The current study sought to investigate the in vitro protein evolution of NDM-1 beta-lactamase using error-prone polymerase chain reaction. Evaluation revealed that variants were not found to confer higher levels of resistance toward meropenem based on amino acid substitutions. Thus, we postulate that increases in transcription or changes in zinc transport may be clinically more relevant to meropenem resistance than amino acid substitutions.